论文部分内容阅读
将人Ｂ７基因亚克隆到逆转录病毒载体ｐＬＸＳＮ中，构建了人Ｂ７逆转录病毒载体ｐＬＸＳＮＢ７，经Ｌｉｐｏｆｅｃｔｉｎ介导转染包装细胞，并通过Ｇ４１８筛选获得较高病毒滴度的细胞株ＰＡ３１７／ｐＬＸＳＮＢ７（３×１０５～１×１０６ｃｆｕ／ｍｌ）。用此逆转录病毒基因转移体系将Ｂ７基因转移到肾癌ＧＲＣ１细胞中，经ＲＴＰＣＲ、间接免疫荧光及ＬＳＡＢ细胞免疫组化等检测方法检测到转基因细胞中Ｂ７ｍＲＮＡ及Ｂ７分子的表达。介导人Ｂ７基因的逆转录病毒基因转移体系的建立，为人肾癌转基因瘤苗的应用研究提供了实验基础。
The human B7 gene was subcloned into the retroviral vector pLXSN to construct a human B7 retroviral vector pLXSNB7, which was transfected into packaging cells via Lipofectin, and cell lines PA317/pLXSNB7 with higher titer were obtained by G418 selection ( 3 x 105 to 1 x 106 cfu/ml). Using this retroviral gene transfer system, B7 gene was transferred to renal cell carcinoma GRC-1 cells. The expression of B7 mRNA and B7 molecules in the transgenic cells was detected by RT-PCR, indirect immunofluorescence and LSAB immunohistochemistry. The establishment of a retroviral gene transfer system that mediates human B7 gene provides an experimental basis for the application research of human kidney cancer transgenic tumor vaccine.